International Journal of Health Sciences and Research www.ijhsr.org ISSN: 2249-9571 Original Research Article A Newly Identified Stain for Mast Cells in Human Mesentery Roopa Ravindranath, R.Victor, Amith Ramos*, V. Sumitra Histology Laboratory, Dept. of Anatomy, St. John’s Medical College, Bangalore-560034. *Correspondence Email: [email protected] Received: 03/01//2013 Revised: 31/01/2013 Accepted: 04/02/2013 ABSTRACT For invitro studies in mesentery toluidine blue and thioninare generally used. We have identified a new stain and found it to be good as it showed good contrast between Mast cells and the background tissue. A simple, cost effective and rapid method for staining mast cells in human mesentery is described. Mesentery was removed from embalmed cadaver and cut into pieces, spread on glass slides, tied with the thread on both sides and fixed in 10% formalin overnight and stored in 70% alcohol. Small pieces of convenient size were cut and transferred into a petri dish, washed thoroughly and was spread on a glass slide. The tissue was stained with Methyl Violet 10B (MV10B) stain for 1 minute, washed, dehydrated, cleared and mounted in DPX. Mast cells were seen against light connective tissue background. Key words: Stain, Mast Cells, Human Mesentery INTRODUCTION can be used to advantage for routine study of Mast cell function has been mast cells. attributed to wound healing, immune allergic reaction and regeneration among MATERIALS AND METHODS other functions. They are widely distributed Mesentery was removed from in the connective tissue. The abundant embalmed cadaver during the routine granules stain metachromatically with dissection of the abdomen. The sheet of thiazide dies such as toluidine blue. They mesentery was cut into small pieces to also stain with copper phthalocyanine dyes accommodate in the slides. The cut pieces such as alcian blue and astra blue. [1] were spread on a slide and tied with thread Astra blue has been used to stain from both sides. The slides were fixed in 10 differentially mast cells in the intestine. % formalin overnight and storedin 70% However the procedure has not been widely alcohol. Small pieces were cut, transferred used because of difficulty in preparing and into a petri dish and were washed using the dye solution. [2] thoroughly. The bits were taken on to the In the present study the contrast slides and stained with MV10 stain for 1 between the deeply stained granules and the minute, washed, dehydrated cleared and lightly stained background facilitates mounted in DPX. detection and counting of mast cells. This International Journal of Health Sciences & Research (www.ijhsr.org) 66 Vol.3; Issue: 2; February 2013 Preparation of the MV10B stain: 2.Wash in water until the color ceases. -MV10B stain 0.25 gms, 3.Dehydrate in absolute alcohol -Autoclave distilled water : 100ml 4.Clear in xylene -Dissolved and filtered with 5.Mount in DPX whatman filter paper and stored in a brown bottle at room temperature. RESULTS Darkly stained granules with light Staining procedure: staining nucleus of mast cells are seen 1.Stain in MV10B stain for one minute against connective tissue background. Fig. 1. Mesentery (x400): Arrows- Mast cells, Fig. 2. Mast Cells in the mesentery (x1000): C- Connective tissue background G- Granules, N- Nucleus DISCUSSION with a variety of mast tissue fixatives makes Thioninhas been used to stain mast it useful for routine use in histopathological cells in mesentery. [3] The methods involves laboratories. staining with a diluted solution of thionin for Methyl violet is the family of organic 30 minutes, differentiated in 0.2 % acetic compound that are mainly used as dyes. acid controlled microscopically and counter Depending on the amount of attached stained with saturated tartrazine in cello methyl groups the colour of the dye can be solve. [4] altered. Its main use is as the purple dye for Previously we had reported a textiles and to give deep violet colours in modified thioninacridine orange stain for paint and ink. Methyl violet 10B is also mast cells in rat mesentery which can be known as crystal violet (and many other completed within 10 minutes and doesn’t names) and has medical uses. require any differentiation. [5] Methyl violet 10B has six methyl The present study involves a new groups. It is known in medicine as gentian single staining technique using which the violet (or crystal violet or pyoctanin(e) & is darkly stained granules stand out in better the active ingredient in gram stain used to contrast on a fairly stained background. The classify bacteria. It is used as a pH indicator simplicity and reproducibility of the with a range between 0 and 1.6. The procedure combined with the compatibility protonated form (found in acidic conditions) International Journal of Health Sciences & Research (www.ijhsr.org) 67 Vol.3; Issue: 2; February 2013 is yellow terming blue violet above pH REFERENCES levels of 1.6. Gentian violet destroys cells 1. Enerback L. Mast cells in and can be used as a disinfectant. heterogeneity – the evolution of the Compounds related to methyl violet are concept of a special mucosal mast potential carcinogens. cell in: Mast cells differentiation and Methyl violet 10B also inhibits the heterogeneity. Bufes (eds) New growth of many gram positive bacteria York, Raven Pres;1986.p.1-16. except streptococci, when used in 2. Bloom G and Kelly JW. The copper conjunction with nalidixic acid (which Pthalocyanin dye “Astra blau” and destroys gram negative bacteria). It can be its staining properties especially the used to isolate the streptococci bacteria for staining of mast cells. Histochem J the diagnosis of an infection. 1960; 2: 48-57 Methyl violet 10B also binds to 3. Cooke HC. A modified thionin DNA. This means it can be used in all technique for mast cells in tissue viability assays in biochemistry. However sections. J of Med. Lab. Tech this binding to DNA will cause replication 1961;18:188 errors in living tissue, possibly leading to 4. Rex N Paul. The use of thionin and mutations and cancer. [6] acridine orange in staining semi thin sections of plant material embedded CONCLUSION in epoxy resin. Stain technology The advantages of the newly 1971;46:195-7 identified MV10B staining techniques are - 5. R Victor, R. Ravindranath et al. A easy to prepare, low dye concentration, no modified thioninacridine orange differentiation, consistent results and easy to stain for mast cells, Indian J Pathol, perform the technique. Microbiol 2004;47(2): 168-169. 6. Methyl Violet [Online]. 2012 Oct 1 ACKNOWLEDGEMENT [cited 2013 Jan 2]; Available from: The secretarial assistance of Mr. Job URL:http:www.en.wikipedia.org/wi Kaayal is acknowledged. ki/Methyl_violet. How to cite this article: Ravindranath R, Victor R, Ramos A et. al. A newly identified stain for mast cells in human mesentery. Int J Health Sci Res. 2013;3(2):66-68. ************************ International Journal of Health Sciences & Research (www.ijhsr.org) 68 Vol.3; Issue: 2; February 2013 .